Different methods for construction of novel binding proteins have been described (Nygren P A and Uhlén M (1997) Curr Opin Struct Biol 7:463-469). One strategy has been to combine library generation and screening with selection for desired properties.
First generation Z variant polypeptides based on a common, first generation scaffold, populations of such molecules and methods involving them have been described in WO95/19374. Additionally, Z variant polypeptides based on a second generation scaffold, populations of such molecules and methods involving them have been described in WO2009/080811. The teachings of these two disclosures are incorporated herein by reference.
For some applications, Z variant polypeptides or populations thereof having improved properties, such as higher alkali stability, low antigenicity, structural stability, amenability to chemical synthesis and hydrophilicity, are desired. WO2009/080811 discloses Z variants having a common scaffold with improved properties, but not every desired property can be obtained by Z variant polypeptides as described therein.
One of the key factors to success for polypeptide pharmaceuticals is their stability. Polypeptides showing a high structural stability will most likely functionally withstand chemical modifications, changes in physical conditions and proteolysis, both during production as well as within the human body. Moreover, stability will influence the active shelf-life of polypeptide pharmaceuticals, as well as the active life of the polypeptide pharmaceutical within the human body.
Hence, there is a continued need for improving the stability of Z variant polypeptides.